Nanoparticles are submicron materials that often possess different properties than bulk material of the same kind. Nanoparticles have been studied for uses in many fields, including diagnostic and therapeutic applications in the life sciences. Because of their small size and unique properties, nanoparticles often have enhanced distribution in the body compared to larger sized particles. Further, nanoparticles may be specifically directed to particular targets in the body by attaching one or more components to the nanoparticle surface (i.e. functionalization). Functionalization of a nanoparticle with a component having affinity for a specific target in the body can direct the nanoparticle to tissues containing the target molecule.
Expansion of nanoparticles at room temperature can facilitate rapid loading of degradation sensitive therapies by passive diffusion. Thermosensitive polymer poly(N-isopropylacrylamide), abbreviated poly(NIPAm), has a physiologically relevant lower critical solution temperature (LCST) between 31-33° C. This LCST causes the polymer to expand at room temperature and contract under physiological conditions. Charged carboxylic co-monomers, such as acrylic acid, are traditionally added to poly(NIPAm) nanoparticles in order to increase colloidal stability and provide a secondary site for chemical modification. In addition to carboxylic co-monomers, it is possible to incorporate sulfated co-monomer 2-acrylamido-2-methyl-1-propanesulfonic acid (AMPS) into poly(NIPAm) nanoparticles.
As described herein, the addition of AMPS into poly(NIPAm) nanoparticles greatly enhances the electrostatic attraction of kinase inhibiting peptides and the nanoparticles which results in increased drug loading. In addition, Applicants have shown that anti-inflammatory kinase inhibiting peptides can be used to target mitogen activated protein kinase activated protein kinase 2 (MK2) to reduce production of pro-inflammatory cytokines and suppress inflammation. As herein described, poly(NIPAm-AMPS) nanoparticles incorporated with or coupled to kinase inhibiting peptides can be used to selectively diffuse through cartilage and significantly suppress cytokine production. For example, the nanoparticle incorporated kinase inhibiting peptides described herein can be used for interarticular therapies, e.g. to treat inflammatory diseases.
Several embodiments of the invention are described by the following enumerated clauses:
1) A composition comprising at least one kinase inhibiting peptide incorporated with a nanoparticle, wherein the nanoparticle comprises at least one copolymerized monomer having an anionic side chain and at least one additional copolymerized monomer.
2) The composition of clause 1 wherein the kinase inhibiting peptide is an MK2 inhibitor peptide.
3) The composition of any one of clauses 1 and 2 wherein the amino acid sequence of the kinase inhibiting peptide has at least 90% sequence identity to a peptide selected from the group consisting of YARAAARQARAKALARQLGVAA, YARAAARQARAKALNRQLGVA, FAKLAARLYRKALARQLGVAA, KAFAKLAARLYRKALARQLGVAA, HRRIKAWLKKIKALARQLGVAA, YARAAARQARAKALNRQLAVAA, and YARAAARQARAKALNRQLAVA.
4) The composition of any one of clause 1 to 3 wherein the at least one copolymerized anionic monomer is 2-acrylamido-2-methyl-1-propanesulfonic acid.
5) The composition of any one of clauses 1 to 4 wherein the at least one additional copolymerized monomer is N-isopropylacrylamide.
6) The composition of any one of clauses 1 to 5 further comprising a crosslinker.
7) The composition of clause 6 wherein the crosslinker is selected from the group consisting of N,O-dimethacryloylhydroxylamine, divinyl adipate, N,N-Bis(acryloyl)cystamine, and N,N′-methylenebisacrylamide.
8) The composition of any one of clauses 1 to 6 further comprising a pharmaceutically acceptable carrier.
9) The composition of clause 8 wherein the carrier is a liquid carrier and is selected from the group consisting of saline, glucose, alcohols, glycols, esters, amides, and a combination thereof.
10) An effective dose of the composition of any of clauses 1 to 9 for administration to a patient, wherein the effective dose ranges from about 1 ng to about 1 mg per kilogram of body weight.
11) An effective dose of the composition of any of clauses 1 to 9 for administration to a patient, wherein the effective dose ranges from about 1 pg to about 10 ng per kilogram of body weight.
12) An effective dose of the composition of any of clauses 1 to 9 for administration to a patient, wherein the effective dose ranges from about 1 μg to about 100 μg per kilogram of body weight.
13) The composition of any one of clauses 1 to 12 wherein the kinase inhibiting peptide has at least 90% sequence identity to YARAAARQARAKALARQLGVAA.
14) The composition of any one of clauses 1 to 12 wherein the kinase inhibiting peptide has at least 90% sequence identity to YARAAARQARAKALNRQLGVA.
15) The composition of any one of clauses 1 to 12 wherein the kinase inhibiting peptide has at least 90% sequence identity to FAKLAARLYRKALARQLGVAA.
16) The composition of any one of clauses 1 to 12 wherein the kinase inhibiting peptide has at least 90% sequence identity to KAFAKLAARLYRKALARQLGVAA.
17) The composition of any one of clauses 1 to 12 wherein the kinase inhibiting peptide has at least 90% sequence identity to HRRIKAWLKKIKALARQLGVAA.
18) The composition of any one of clauses 1 to 12 wherein the kinase inhibiting peptide has at least 90% sequence identity to YARAAARQARAKALNRQLAVAA.
19) The composition of any one of clauses 1 to 12 wherein the kinase inhibiting peptide has at least 90% sequence identity to YARAAARQARAKALNRQLAVA.
20) The composition of any one of clauses 1 to 19 wherein the at least one copolymerized anionic monomer is 2-acrylamido-2-methyl-1-propanesulfonic acid, and wherein the at least one additional copolymerized monomer is N-isopropylacrylamide.
21) The composition of any one of clauses 1 to 20 wherein the at least one copolymerized anionic monomer is 2-acrylamido-2-methyl-1-propanesulfonic acid, wherein the at least one additional copolymerized monomer is N-isopropylacrylamide, and wherein the kinase inhibiting peptide is KAFAKLAARLYRKALARQLGVAA.
22) The composition of any one of clauses 1 to 21 wherein the at least one copolymerized anionic monomer is 2-acrylamido-2-methyl-1-propanesulfonic acid, wherein the at least one additional copolymerized monomer is N-isopropylacrylamide, wherein the crosslinker is N,O-dimethacryloylhydroxylamine, and wherein the kinase inhibiting peptide is KAFAKLAARLYRKALARQLGVAA.
23) The composition of any one of clauses 1 to 22 wherein the at least one additional copolymerized monomer and the copolymerized anionic monomer are present in the composition at a ratio selected from the group consisting of about 5:1, about 8:1, about 9:1, about 10:1, about 11:1, about 12:1, and about 15:1.
24) The composition of any one of clauses 1 to 23 wherein the at least one additional copolymerized monomer and the copolymerized anionic monomer are present in the composition at a ratio of about 10:1.
25) A method for treating osteoarthritis, the method comprising the step of administering a composition comprising at least one kinase inhibiting peptide incorporated with a nanoparticle, wherein the nanoparticle comprises at least one copolymerized anionic monomer and at least one additional copolymerized monomer.
26) The method of clause 25 wherein the kinase inhibiting peptide is an MK2 inhibitor peptide.
27) The method of any one of clauses 25 and 26 wherein the amino acid sequence of the kinase inhibiting peptide has at least 90% sequence identity to a peptide selected from the group consisting of YARAAARQARAKALARQLGVAA, YARAAARQARAKALNRQLGVA, FAKLAARLYRKALARQLGVAA, KAFAKLAARLYRKALARQLGVAA, HRRIKAWLKKIKALARQLGVAA, YARAAARQARAKALNRQLAVAA, and YARAAARQARAKALNRQLAVA.
28) The method of any one of clause 25 to 27 wherein the at least one copolymerized sulfonic monomer is 2-acrylamido-2-methyl-1-propanesulfonic acid.
29) The method of any one of clauses 25 to 28 wherein the at least one additional copolymerized monomer is N-isopropylacrylamide.
30) The method of any one of clauses 25 to 29 wherein the nanoparticle further comprising a crosslinker.
31) The method of clause 30 wherein the crosslinker is selected from the group consisting of N,O-dimethacryloylhydroxylamine, divinyl adipate, N,N-Bis(acryloyl)cystamine, and N,N′-methylenebisacrylamide.
32) The method of any one of clauses 25 to 31 further comprising a pharmaceutically acceptable carrier.
33) The method of clause 32 wherein the carrier is a liquid carrier and is selected from the group consisting of saline, glucose, alcohols, glycols, esters, amides, and a combination thereof.
34) The method of any one of clauses 25 to 33 wherein the composition is administered to a patient in an effective dose, wherein the effective dose ranges from about 1 ng to about 1 mg per kilogram of body weight.
35) The method of any one of clauses 25 to 33 wherein the composition is administered to a patient in an effective dose, wherein the effective dose ranges from about 1 pg to about 10 ng per kilogram of body weight.
36) The method of any one of clauses 25 to 33 wherein the composition is administered to a patient in an effective dose, wherein the effective dose ranges from about 1 μg to about 100 μg per kilogram of body weight.
37) The method of any one of clauses 25 to 33 wherein the composition is administered to a patient in an effective dose, of any one of clauses 25 to 36 wherein the kinase inhibiting peptide has at least 90% sequence identity to YARAAARQARAKALARQLGVAA.
38) The method of any one of clauses 25 to 33 wherein the kinase inhibiting peptide has at least 90% sequence identity to YARAAARQARAKALNRQLGVA.
39) The method of any one of clauses 25 to 33 wherein the kinase inhibiting peptide has at least 90% sequence identity to FAKLAARLYRKALARQLGVAA.
40) The method of any one of clauses 25 to 33 wherein the kinase inhibiting peptide has at least 90% sequence identity to KAFAKLAARLYRKALARQLGVAA.
41) The method of any one of clauses 25 to 33 wherein the kinase inhibiting peptide has at least 90% sequence identity to HRRIKAWLKKIKALARQLGVAA.
42) The method of any one of clauses 25 to 33 wherein the kinase inhibiting peptide has at least 90% sequence identity to YARAAARQARAKALNRQLAVAA.
43) The method of any one of clauses 25 to 33 wherein the kinase inhibiting peptide has at least 90% sequence identity to YARAAARQARAKALNRQLAVA.
44) The method of any one of clauses 25 to 43 wherein the at least one copolymerized anionic monomer is 2-acrylamido-2-methyl-1-propanesulfonic acid, and wherein the at least one additional copolymerized monomer is N-isopropylacrylamide.
45) The method of any one of clauses 25 to 44 wherein the at least one copolymerized anionic monomer is 2-acrylamido-2-methyl-1-propanesulfonic acid, wherein the at least one additional copolymerized monomer is N-isopropylacrylamide, and wherein the kinase inhibiting peptide is KAFAKLAARLYRKALARQLGVAA.
46) The method of any one of clauses 25 to 45 wherein the at least one copolymerized anionic monomer is 2-acrylamido-2-methyl-1-propanesulfonic acid, wherein the at least one additional copolymerized monomer is N-isopropylacrylamide, wherein the crosslinker is N,O-dimethacryloylhydroxylamine, and wherein the kinase inhibiting peptide is KAFAKLAARLYRKALARQLGVAA.
47) The method of any one of clauses 25 to 46 wherein the at least one additional copolymerized monomer and the copolymerized anionic monomer are present in the composition at a ratio selected from the group consisting of about 5:1, about 8:1, about 9:1, about 10:1, about 11:1, about 12:1, and about 15:1.
48) The method of any one of clauses 25 to 47 wherein the at least one additional copolymerized monomer and the copolymerized anionic monomer are present in the composition at a ratio of about 10:1.
49) The composition or method of any one of the preceding clauses wherein the anionic monomer is a sulfonic monomer.
50) The composition or method of any one of the preceding clauses wherein the composition is a pharmaceutical composition.